Phagocytosis of crocidolite asbestos induces oxidative stress, DNA damage, and apoptosis in mesothelial cells

Our plain-English summary of Phagocytosis of crocidolite asbestos induces oxidative stress, DNA damage, and apoptosis in mesothelial cells, by W. Liu, J.D. Ernst, V.C. Broaddus (American Journal of Respiratory Cell and Molecular Biology, 2000).

The study

• Authors: W. Liu, J.D. Ernst, V.C. Broaddus
• Journal: American Journal of Respiratory Cell and Molecular Biology
• Published: 2000;23(3):371-8
• Cited by: 74 articles, as counted by NIH iCite
• Read it: PubMed · Publisher

Abstract

Phagocytosis of asbestos fibers may be a necessary step for asbestos-induced injury to mesothelial cells, but this has not been established because quantification of fiber uptake is difficult and ways to increase fiber phagocytosis without also increasing total dose were not available. We quantified phagocytosis by counting intracellular fibers after removing adherent fibers with trypsin; we selectively increased fiber phagocytosis by coating crocidolite asbestos fibers with the adhesive serum protein vitronectin (VN), which we have shown increases fiber uptake via integrins. We measured various aspects of asbestos-induced cytotoxicity: intracellular oxidation by the shift of fluorescence of cells loaded with an oxidative probe, DNA strand breakage by the alkaline unwinding ethidium bromide fluorometric assay, apoptosis by annexin V binding and by nuclear morphology, and cell-cycle progression. We found that, compared with control fibers or particles, asbestos increased intracellular oxidation, DNA strand breakage, and apoptosis. Selective increases in fiber uptake by VN-coating of the fibers further increased the oxidation, DNA strand breakage, and apoptosis, and induced a cell-cycle arrest in G2/M. Selective decreases in fiber uptake by cytochalasin or by integrin blockade with RGD peptides inhibited several of these measures of injury. We conclude that phagocytosis is important and perhaps necessary for asbestos-induced injury to mesothelial cells.

In plain English

This was a laboratory study of rabbit pleural mesothelial cells, not of people. The authors asked whether cells must take in crocidolite asbestos fibers to be harmed by them. They increased uptake by coating fibers with vitronectin, a blood protein. Cytochalasin B reduced uptake of all fibers. RGD peptides, at the concentration chosen, reduced uptake of vitronectin-coated fibers but not uncoated ones. Wollastonite fibers and glass beads served as comparisons. After 4 hours, 64 ± 7% of cells exposed to vitronectin-coated fibers held more than four fibers, against 43 ± 5% with uncoated fibers. Asbestos raised markers of oxidative stress and DNA strand breaks, and coated fibers raised them further. Wollastonite caused no significant DNA breakage. Glass beads were taken in but did not shift the fluorescent oxidation probe. Cytochalasin completely blocked asbestos-related DNA damage. Coated fibers also increased apoptosis (a form of cell death) and briefly held up cells at one stage of cell division. The authors concluded that, at least over the duration of their experiments, fiber uptake was necessary for asbestos to injure these cells.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: laboratory (in vitro) cell-culture experiment
• Who was studied: Primary rabbit pleural mesothelial cells in culture, used between passages 3 and 7; results come from 3 to 10 repeated experiments depending on the assay
• Exposure: Crocidolite asbestos fibers, either uncoated or coated with vitronectin (VN), bovine serum albumin (BSA) or fibronectin (FN). Comparison particles were wollastonite (a calcium silicate fiber) and glass beads. Uptake was reduced with cytochalasin B or RGD peptides. Exposures lasted 4 to 24 hours.
• Main results:
  • Fiber uptake: 64 ± 7% of cells exposed to VN-coated fibers had more than four internal fibers, compared with 43 ± 5% of cells exposed to uncoated fibers (four experiments). Cytochalasin reduced uptake to 10 ± 3% for uncoated and 20 ± 7% for VN-coated fibers.
  • At the concentration chosen for the experiments, RGD peptides reduced uptake of VN-coated fibers (47 ± 4% of cells) but not of uncoated fibers (45 ± 6%).
  • At higher concentrations, RGD peptides made the cells round up, which reduced uptake of all fibers. These data were not shown.
  • In the oxidation assay, VN-coated crocidolite caused a greater oxidative shift in the cells than BSA-coated crocidolite.
  • Glass beads caused no shift in the fluorescent oxidation probe, even though cells took them up.
  • After 4 hours, VN-coated crocidolite caused more DNA strand breakage than uncoated or BSA-coated crocidolite.
  • Wollastonite, the comparison fiber, did not cause significant DNA strand breakage, whether coated with VN or uncoated.
  • DMSO, a hydroxyl radical scavenger, blocked the DNA damage caused by both uncoated and VN-coated crocidolite.
  • Cytochalasin completely blocked the DNA damage caused by uncoated, BSA-coated and VN-coated crocidolite.
  • RGD peptides blocked DNA damage only from VN-coated fibers. Neither blocker affected DNA damage caused by hydrogen peroxide.
  • Late apoptosis or necrosis (10 experiments): 9.4 ± 0.6% of cells exposed to VN-coated crocidolite, compared with 5.4 ± 0.6% for uncoated or other protein-coated crocidolite and 2.9 ± 0.3% with no fiber.
  • Apoptosis judged by nuclear appearance (3 experiments): 14.0 ± 1.0% of cells exposed to VN-coated asbestos, compared with 11.0 ± 2.0% for uncoated asbestos and 4.0 ± 1.0% with no fiber.
  • RGD peptides, but not the control RGE peptides, inhibited apoptosis caused by VN-coated crocidolite.
  • At 12 hours, more cells exposed to VN-coated fibers were in the G2/M phase of the cell cycle. By 24 hours, this difference was no longer statistically significant.
• The authors' conclusion: The authors concluded that, at least over the duration of these experiments, phagocytosis (uptake) of asbestos fibers is necessary for asbestos-induced cytotoxicity to mesothelial cells. In the abstract they described phagocytosis as important and perhaps necessary.
• Limitations the authors state:
  • The authors could not assess whether fibers stuck to the cell surface contributed to the oxidation signal, because cells were detached before that assay, which removed those fibers.
  • Cytochalasin could not be used in the apoptosis experiments because it caused apoptosis by itself.
  • The authors attributed their failure to show a G2/M delay with uncoated asbestos to the low-serum conditions they used.

Topics

The National Library of Medicine indexes this study under subjects our site covers: Crocidolite.

Medical terms in this study, explained in our Lexicon: Phagocytosis, Crocidolite, Oxidative Stress, Apoptosis, Mesothelial Cells, Vitronectin.

Related studies

• Vitronectin adsorption to chrysotile asbestos increases fiber phagocytosis and toxicity for mesothelial cells — J. Wu, W. Liu, et al. (2000)
• Asbestos induces mitochondrial DNA damage and dysfunction linked to the development of apoptosis — A. Shukla, M. Jung, et al. (2003)
• Potential role of ferritin heavy chain in oxidative stress and apoptosis in human mesothelial and mesothelioma cells: implications for asbestos-induced oncogenesis — W. Aung, S. Hasegawa, et al. (2007)
• Patterns of 8-hydroxydeoxyguanosine formation in DNA and indications of oxidative stress in rat and human pleural mesothelial cells after exposure to crocidolite asbestos — H. Fung, Y.W. Kow, et al. (1997)
• Cisplatin and resveratrol induce apoptosis and autophagy following oxidative stress in malignant mesothelioma cells — Y.J. Lee, G.J. Lee, et al. (2016)
• Detection of 8-oxo-2'-deoxyguanosine, a marker of oxidative DNA damage, in culture medium from human mesothelial cells exposed to crocidolite asbestos — Q. Chen, J. Marsh, et al. (1996)

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